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Nuclear targeting of Akt enhances kinase activity and survival of cardiomyocytes

Akt 的核靶向增强激酶活性和心肌细胞的存活

基本信息

批准号：
14570760
负责人：
SHIRAISHI Isao
金额：
$ 1.92万
依托单位：
Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14570760/
关键词：
Heart Failure Gene Therapy Apoptosis Akt Nuclear Localization signal 細胞内シグナル伝達核移行

项目摘要

Heart failure is associated with death of cardiomyocytes leading to loss of contractility. Previous studies using membrane-targeted Akt (myristolated-Akt), an enzyme involved in anti-apoptotic signaling, showed inhibition of cell death and prevention of pathogenesis induced by cardiomyopathic stimuli. However, recent studies by our group have found accumulation of activated Akt in the nucleus, suggesting that biologically relevant target(s) of Akt activity may be located there. To test this hypothesis, a targeted Akt construct was created to determine the anti-apoptotic action of nuclear Akt accumulation. Nuclear localization of the adenovirally-encoded Akt construct was confirmed by confocal microscopy. Cardiomyocytes expressing nuclear targeted Akt showed no evidence of morphologic remodeling such as altered myofibril density or hypertrophy. Nuclear targeted Akt significantly elevated levels of phospho-Akt and kinase activity and inhibited apoptosis as effectively as myristolated-Akt using in hypoxia-induced cell death. Transgenic overexpression of nuclear targeted Akt did not result in hypertrophic remodeling, altered cardiomyocyte DNA content or nucleation, or enhanced phosphorylation of typical cytoplasmic Akt substrates, yet transgenic hearts were protected from ischemia-reperfusion injury. Gene array analyses demonstrated changes in the transcriptional profile of Akt/nuc hearts compared to nontransgenic controls distinct from prior characterizations of Akt expression in transgenic hearts. Collectively, these experiments show that targeting of Akt to the nucleus mediates inhibition of apoptosis without hypertrophic remodeling, opening new possibilities for therapeutic applications of nuclear targeted Akt to inhibit cell death associated with heart disease.

心力衰竭与心肌细胞死亡相关，导致收缩力丧失。先前的研究表明，利用膜靶向Akt (myristolate -Akt)，一种参与抗凋亡信号传导的酶，可以抑制心肌病刺激诱导的细胞死亡和预防发病机制。然而，我们小组最近的研究发现活化的Akt在细胞核中积累，这表明Akt活性的生物学相关靶点可能位于那里。为了验证这一假设，我们构建了靶向Akt构建体，以确定核Akt积累的抗凋亡作用。共聚焦显微镜证实了腺病毒编码的Akt结构的核定位。表达核靶向Akt的心肌细胞未显示肌原纤维密度改变或肥大等形态学重构的证据。在缺氧诱导的细胞死亡中，核靶向Akt显著提高磷酸化Akt水平和激酶活性，抑制细胞凋亡的效果与肉芽化Akt相同。转基因过表达核靶向Akt不会导致肥厚重塑、心肌细胞DNA含量或成核改变，也不会增强典型细胞质Akt底物的磷酸化，但转基因心脏可免受缺血-再灌注损伤。基因阵列分析显示，与非转基因对照相比，Akt/nuc心脏的转录谱发生了变化，这与先前转基因心脏中Akt表达的特征不同。综上所述，这些实验表明，Akt靶向细胞核介导细胞凋亡的抑制，而没有肥厚重塑，这为核靶向Akt抑制心脏病相关细胞死亡的治疗应用开辟了新的可能性。